Fibrosis, the hallmark of systemic sclerosis (SSc), represents the transformation of normal wound-healing into a deregulated self-sustaining process. Recent evidence indicates that in SSc injury results in accumulation of tissue breakdown products that can be recognized as danger signals by innate immune sensors. We showed that fibroblasts express and can be activated through toll-like receptors (TLRs) in much the same way that immune cells recognize microbial pathogen-associated patterns. Furthermore,tissue expression of TLR4, and its putative endogenous ligand hyaluronic acid, are both elevated in SSc. We hpothesize that accumulation of endogenous TLR ligands in injured tissue in SSc drives TLR-mediated amplification and persistence of fibroblast activation, resulting in intractable fibrosis. In this proposal we will examine the expression, regulation and function of fibroblast TLRs in SSc. In Specific Aim 1, we will evaluate cellular TLR4 and endogenous TLR ligand levels in SSc, define a cutaneous TLR signature, and correlate the tissue TLR signature with clinical parameters in SSc patients. In Specific Aim 2 we will examine the regulation of TLR expression, and the mechanism of TLR4-dependent fibroblast activation. In Specific Aim 3 we will examine the role of TLR4 and endogenous TLR4 ligands in scleroderma models using loss-of-function approach, and in Specific Aim 4, we will examine if fibroblast-restricted TLR overexpression can induce the transition of self-limited healing into progressive intractable fibrogenesis. These studies will provide the first insight into the role of fibroblast TLR signaling in the pathogenesis of SSc. The proposed research will accelerate the development of novel anti-fibrotic treatments for SSc. PUBLIC HEALTH RELEVANCE: Systemic sclerosis resembles uncontrolled wound healing, where healing occurs by fibrosis rather than tissue regeneration. The factors responsible for loss of regulatory mechanisms normally controlling wound healing are unknown. We propose that innate immune sensors on lesional fibroblasts are activated within the damaged tissue environment, transforming self-limited repair into intractable fibrosis. These studies will test this hypothesis. The results may elucidate novel strategies for the treatment of SSc.